Successful vaccination against HIV will likely require the generation of coordinated humoral and cellular immunity. Among current strategies, protein vaccines fail to stimulate cellular responses, while plasmid DNA or live recombinant vectors capable of promoting cellular and humoral responses together face other issues, such as potency in humans (DNA), or challenges of vector-specific immunity and/or safety (live vectors). We propose here a new class of lipid-based vesicles for protein vaccine delivery, composed of multilamellar lipid vesicles stabilized by the introduction of crosslinks connecting the bilayers of the structure. These novel materials are synthesized and encapsulate protein antigens in a mild all-aqueous process, favoring the preservation of epitopes on complex antigens. In preliminary studies, we show that these novel vaccine carriers can be repeatedly administered for homologous boosting: Following a prime and 2 booster immunizations with a model antigen, these particles massively expand antigen-specific CD8+ T-cells, to the best of our knowledge substantially stronger than any previously reported protein vaccine. These T-cells were functional in terms of their capacity to produce IFN-?, and formed a substantial memory population a month after the second boost. In other preliminary studies, we found that this same system drives potent humoral responses against a candidate malaria antigen. Based on these promising initial results, we propose here to test the promise of this new vaccine delivery approach in the context of HIV vaccination, aiming to determine whether potent, durable T-cell and humoral responses can be elicited against HIV gag and env antigens. In addition, we will explore whether particular aspects of this vaccine system (e.g., the deposition of particles at the injection site) can be exploited to direct the localization of memory cells to key mucosal tissue sites that could promote protection against HIV infection. Our specific aims are: (1) How do the humoral and cellular responses to gag and env antigens evolve as a function of antigen dose and boosting with ICMV particle vaccination? (2) Are alternative clinically-relevant TLR agonist/danger signals capable of eliciting stronger immune responses than MPLA when delivered by ICMV vectors? (3) How do particles retained at the injection site influence memory/effector cell trafficking following ICMV particle immunization, compared to traditional protein immunization? (4) Can microneedle delivery be used to allow repeated needle-free particle boosting to be performed, without the need for refrigerated vaccines? Identification of ICMV formulations that can raise immune responses against HIV immunogens comparable to those shown in our preliminary data for OVA or malaria antigens will set the stage for subsequent testing in macaque protection models (either using ICMV vaccination alone or in tandem with other strong HIV vaccine candidates. PUBLIC HEALTH RELEVANCE: In this application, we propose research aimed at testing the promise of a new class of lipid-based nanoparticles for vaccination against HIV. These studies will shed important new light on the mechanisms of immune stimulation by this nanoparticle vaccine, and if successful, these materials will provide a powerful hew vaccine complementary to existing platforms being tested in the effort to develop a protective AIDS vaccine.